1) Beagle dogs vaccinated once intradermally (i.d.) in each ear pinna with a rabies DNA vaccine expressing the rabies virus G produced elevated levels of neutralizing antibody that persist for more than one year. In contrast, dogs vaccinated with this DNA vaccine in the quadriceps muscle, via gene gun in the ear pinnae, or i.d. in the neck elicit minimal or undetectable levels of neutralizing antibody.2) Beagle dogs vaccinated once i.d. in ear pinnae one year prior to infection are protected 100% against a lethal challenge of a salivary gland homogenate obtained from a rabid dog naturally infected with a coyote rabies virus variant. Dogs vaccinated once i.m. with the DNA vaccine are not protected. 3)Persistent infectious rabies virus and/or rabies viral RNA are present in mice 20>500 days after virus injection. Virus and/or genome are primarily detected in blood, bone marrow, injected muscle, and the brain/spinal cord. Rabies virus genome persists in tissues of 30% of immunodeficient mice injected with an avirulent rabies virus. The sensitivity for detecting viral RNA in persistently-infected mice increases 100-fold using real-time Taq-Man PCR analysis. 4)Viremia studies using Taq-Man PCR analysis to detect viral RNA show that there are 2 phases of viremia in mice experimentally infected intramuscularly with rabies virus. The initial phase occurs immediately and up to 2 days after virus injection. After that time (3-30 days), viral RNA is not detected in the blood of mice that remain free of clinical disease. More importantly, a second viremia occurs in mice that develop clinical disease and are exsanguinated 2-4 days after onset of paralysis. 5) Phosphorothioate oligonucleotides (PS-ODNs) are potent inhibitors of in vitro rabies virus replication. The antiviral activity increases with the length of the PS-ODN (40mer>20mer>10mer), and is not dependent on a specific oligonucleotide sequence. Experiments are in progress to determine the in vivo antiviral activity of the PS-ODNs.